Abstract The PI3K/AKT/mTOR axis is one of the most commonly mutated pathways in cancer, where aberrant activation promotes cell growth, proliferation, and survival. However, selective inhibitors targeting PI3K/AKT/mTOR have been hindered by their inability to effectively induce cell death in certain cancers. With the introduction of inhibitors that directly inhibit key pro-survival factors such as BCL-2 and BCL-XL (ABT-263 and ABT-199), the potential to achieve cancer cell death using combinations of targeted inhibitors has become a realizable goal. Here we show that selective inhibition of key components in the PI3K pathway (PI3K, AKT, or mTOR) potently sensitized a panel of DLBCL cell lines to ABT-263-induced apoptosis. While the degree of sensitization varied according to which PI3K pathway component was targeted, dual inhibition of both PI3K and mTOR consistently elicited the most potent sensitization across several cell lines. Previous work in other cancer types has linked the potency of this combination to the capacity of PI3K pathway inhibitors to reduce MCL-1 in an mTORC1-dependent manner. However, we found that this was not the case for DLBCL. For example, ABT-263 resistance induced by over-expression of MCL-1 was overcome when cells were co-treated with a dual-PI3K/mTOR inhibitor NVP-BEZ235 despite maintained expression of MCL-1. Instead, inhibition of the PI3K pathway led to a general increase in mitochondrial priming as measured by BH3 profiling. This occurred through distinct effects from both AKT and mTORC1 on the abundance of multiple BCL-2 family proteins at the mitochondria including reductions in the pro-survival proteins BCL-2, MCL-1, and BCL-XL as well as increases in Bim and Bad. In addition to these direct effects on BCL-2 family proteins, we found that inhibition of the PI3K pathway reduced expression of cancer-relevant proteins c-Myc and eIF4G as determined by reverse phase protein array. Thus, this project highlights the broad effects of dual-PI3K/mTOR inhibitors that sensitize DLBCL to apoptosis, and suggests that inhibition of the PI3K pathway has distinct effects on cell survival signaling among different cancer cell types. Citation Format: Jong-Hoon Scott Lee, Sarah Tang, Veronica Ortiz, David A. Fruman. Vertical inhibition of the PI3K pathway potently sensitizes diffuse large B cell lymphoma to BCL-2 antagonism. [abstract]. In: Proceedings of the AACR Special Conference: Targeting the PI3K-mTOR Network in Cancer; Sep 14-17, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(7 Suppl):Abstract nr B45.

Mechanistic target of rapamycin (mTOR) is a serine/threonine kinase that functions as a key regulator of cell growth, division and survival. Many haematologic malignancies exhibit elevated or aberrant mTOR activation, supporting the launch of numerous clinical trials aimed at evaluating the potential of single agent mTOR-targeted therapies. While promising early clinical data using allosteric mTOR inhibitors (rapamycin and its derivatives, rapalogs) have suggested activity in a subset of haematologic malignancies, these agents have shown limited efficacy in most contexts. Whether the efficacy of these partial mTOR inhibitors might be enhanced by more complete target inhibition is being actively addressed with second generation ATP-competitive mTOR kinase inhibitors (TOR-KIs), which have only recently entered clinical trials. However, emerging preclinical data suggest that despite their biochemical advantage over rapalogs, TOR-KIs may retain a primarily cytostatic response. Rather, combinations of mTOR inhibition with other targeted therapies have demonstrated promising efficacy in several preclinical models. This review investigates the current status of rapalogs and TOR-KIs in B cell malignancies, with an emphasis on emerging preclinical evidence of synergistic combinations involving mTOR inhibition.

Abstract Targeting mTOR signaling is a promising approach for treating blood cancers. We reported that mTOR kinase inhibitors, including PP242 and MLN0128, synergize with ABL tyrosine kinase inhibitors (TKIs) to cause cell cycle arrest and death in acute leukemia cells driven by BCR-ABL. mTOR kinase inhibitors are more effective than rapamycin in these models and have minimal effects on normal hematopoietic cells and immune responses at anti-leukemic doses. Ongoing studies indicate that mTOR kinase inhibitors are immunosuppressive at slightly higher concentrations. Moreover, the compounds are generally not cytotoxic as single agents in leukemia or lymphoma models. These findings emphasize the need for rational combinations to unleash the therapeutic potential of mTOR kinase inhibitors. To this end, we have tested various classes of anti-cancer agents guided by gene expression and proteomic data. Our data reveal synergies between mTOR kinase inhibitors with histone deacetylase inhibitors in B-ALL, and with BCL2 antagonists in DLBCL. Unexpectedly, mTOR kinase inhibitors protect B-ALL cells from methotrexate and 6-mercaptopurine, chemotherapeutic agents used in the treatment of B-ALL patients. ABL TKIs can also protect B-ALL cells from methotrexate by inhibiting downstream mTOR signaling. Together these studies identify potential applications and limitations of mTOR-targeted therapy in blood cancers. Citation Format: Thanh-Trang Vo, Jong-Hoon Scott Lee, Lomon So, Brandon Beagle, Matthew R. Janes, David A. Fruman. Mechanisms of resistance to mTOR inhibitors in leukemia and lymphoma. [abstract]. In: Proceedings of the AACR Special Conference: Targeting the PI3K-mTOR Network in Cancer; Sep 14-17, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(7 Suppl):Abstract nr IA16.

The mechanistic target of rapamycin (mTOR) is a kinase whose activation is associated with poor prognosis in pre-B cell acute lymphoblastic leukemia (B-ALL). These and other findings have prompted diverse strategies for targeting mTOR signaling in B-ALL and other B-cell malignancies. In cellular models of Philadelphia Chromosome-positive (Ph+) B-ALL, mTOR kinase inhibitors (TOR-KIs) that inhibit both mTOR-complex-1 (mTORC1) and mTOR-complex-2 (mTORC2) enhance the cytotoxicity of tyrosine kinase inhibitors (TKIs) such as dasatinib. However, TOR-KIs have not shown substantial efficacy at tolerated doses in blood cancer clinical trials. Selective inhibition of mTORC1 or downstream effectors provides alternative strategies that may improve selectivity towards leukemia cells. Of particular interest is the eukaryotic initiation factor 4F (eIF4F) complex that mediates cap-dependent translation. Here we use novel chemical and genetic approaches to show that selective targeting of either mTORC1 kinase activity or components of the eIF4F complex sensitizes murine BCR-ABL-dependent pre-B leukemia cells to dasatinib. SBI-756, a small molecule inhibitor of eIF4F assembly, sensitizes human Ph+ and Ph-like B-ALL cells to dasatinib cytotoxicity without affecting survival of T lymphocytes or natural killer cells. These findings support the further evaluation of eIF4F-targeted molecules in combination therapies with TKIs in B-ALL and other blood cancers.